Simultaneous silencing of TGF-β1 and COX-2 reduces human skin hypertrophic scar through activation of fibroblast apoptosis

// Jia Zhou 1, * , Yixuan Zhao 1, * , Vera Simonenko 2, * , John J. Xu 3 , Kai Liu 1 , Deling Wang 3 , Jingli Shi 3 , Tianyi Zhong 3 , Lixia Zhang 3 , Lun Zeng 4 , Bin Huang 4 , Shenggao Tang 5 , Alan Y. Lu 5 , A. James Mixson 6 , Yangbai Sun 1 , Patrick Y. Lu 2 and Qingfeng Li 1 1 Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China 2 Sirnaomics, Inc., Gaithersburg, MD, USA 3 Suzhou Sirnaomics Pharmaceuticals, Ltd., Biobay, Suzhou, China 4 Guangzhou Xiangxue Pharmaceuticals, Co. Ltd., Guangzhou, China 5 Guangzhou Nanotides Pharmaceuticals, Co. Ltd., Guangzhou, China 6 Department of Pathology, School of Medicine, University of Maryland, Baltimore, MD, USA * These authors contributed equally to this work and share co-first authorship Correspondence to: Qingfeng Li, email: dr.liqingfeng@shsmu.edu.cn Patrick Y. Lu, email: plu@sirnaomics.com Yangbai Sun, email: drsunyb@foxmail.com Keywords: siRNA therapeutics, hypertrophic scar, TGF-β1, COX-2, synergistic effect Received: November 16, 2016 Accepted: August 26, 2017 Published: September 14, 2017 ABSTRACT Excessive skin scars due to elective operations or trauma represent a challenging clinical problem. Pathophysiology of hypertrophic scars entails a prolonged inflammatory and proliferative phase of wound healing. Over expression of TGF-β1 and COX-2 play key regulatory roles of the aberrant fibrogenic responses and proinflammatory mediators. When we silenced TGF-β1 and COX-2 expression simultaneously in primary human fibroblasts, a marked increase in the apoptotic cell population occurred in contrast to those only treated with either TGF-β1 or COX-2 siRNA alone. Furthermore, using human hypertrophic scar and skin graft implant models in mice, we observed significant size reductions of the implanted tissues following intra-scar administration of TGF-β1/COX-2 specific siRNA combination packaged with Histidine Lysine Polymer (HKP). Gene expression analyses of those treated tissues revealed silencing of the target gene along with down regulations of pro-fibrotic factors such as α-SMA, hydroxyproline acid, Collagen 1 and Collagen 3. Using TUNEL assay detection, we found that the human fibroblasts in the implanted tissues treated with the TGF-β1/COX-2 siRNAs combination exhibited significant apoptotic activity. Therefore we conclude that a synergistic effect of the TGF-β1/COX-2siRNAs combination contributed to the size reductions of the hypertrophic scar implants, through activation of fibroblast apoptosis and re-balancing between scar tissue deposition and degradation.